Anti-tumor composition

ABSTRACT

The present invention provides composition having as active ingredients a stilbene derivative and a platinum coordination compound which is highly efficacious and highly safe for treating tumors, particularly for the treatment of solid or malignant tumors and thus methods of cancer and tumor treatment using the composition are also provided.

CONTINUING DATA

This application is a continuation of PCT/JP99/01633, filed Mar. 29,1999, the entire contents of which are herein incorporated by reference.

BACKGROUND OF THE INVENTION

1. Field of the Invention

This invention relates to a novel antitumor or anti-neoplastic agent,composition and treatment. More particularly, it is directed tocompositions comprising a stilbene derivative and a platinumcoordination compound, such as the drug Cisplatin. Such compositionshave been discovered to be safe and highly effective anti-neoplasticagents and are thus useful for the prevention, amelioration, treatmentor cure of neoplasms, cancer, and tumors, particularly malignant orsolid tumors. These compositions may optionally include othertherapeutic compounds or agents. The invention is also directed tomethods of using a composition comprising a stilbene derivative and aplatinum coordination compound, and to methods of co-administration ofthese compounds to prevent, ameliorate, treat, or cure neoplasms, canceror tumors.

2. Discussion of the Background

There is a pronounced need for safe and more efficacious anti-tumoragents. While a wide variety of chemotherapeutic agents are presentlyused for the treatment, suppression and prevention of tumors, tumors maydevelop a resistance to such agents, especially highly malignant orsolid tumors. Thus, tumor relapse is a common problem. Also, existingagents, even if effective, may be inconvenient to administer ineffective dosages and have inadequate therapeutic indexes. Thus,patients may suffer from pain and other side-effects of theiradministration, especially from the administration of high doses ofanti-tumor agents with relatively low potencies.

Platinum coordination compounds, such as Cisplatin and otherdiamino-platinum complexes, have been widely used in humans aschemotherapeutic drugs. The platinum coordination compound used in thepresent invention is such a compound which gives platinum preferably inan ionic form, and preferably is a compound exhibiting substantialanti-tumor activity, more preferably a platinum coordination compoundexhibiting tumor cell proliferation preventative or inhibitingproperties. Many platinum coordination compounds used in the presentinvention are commercially available or may be manufactured by known orroutinely used techniques. However, these compounds are nottherapeutically effective for all patients or all types of tumors. Forinstance, numerous attempts have been made to improve Cisplatin-basedtherapy by using Cisplatin in combination with other drugs. Althoughsome attempts have achieved improvements in efficacy, these compositionswere not sufficiently effective in treating tumors, reducing tumorburden, or in relieving the pain and other complications suffered bytumor patients. Thus, there is a need to discover other agents whichtogether with a platinum coordination compound would be more efficaciousin treating tumors.

Cisplatin or cis-dichlorodiammine platinum (II) has been successfullyused for long as a chemotherapeutic drug in the therapy of variousmalignant tumors in the human being.

More recently, other diamino-platinum complexes have shown efficacy aschemotherapeutic drugs in curing various malignant tumors in the humanbeing. Examples of the diamino-platinum complexes may be spiroplatinumand carboplatinum.

Cisplatin and other diamino-platinum complexes have been widely used aschemotherapeutic drugs in the humans. However, these are nottherapeutically efficient for all patients or all sorts of tumors. Inexpectation of possibly increasing therapeutic efficacy, numerousattempts have been made towards using Cisplatin in combination withVindesine, see Garalla, R. J. et al., Ann. Intern. Med. 95: 414-420(1980) or using Cisplatin in combination with VP-16, see Congeval, E. etal., Cancer, 51: 2751-2756 (1982). Although such combined applicationhas achieved certain improvement in the efficacy ratio, it cannot besaid that the tumor burden or pains of tumor-bearing patients has beencompletely relieved by these measures.

Stilbene derivatives which have cis-stilbene as a fundamental skeleton,are known to strongly inhibit mitosis and cause cytotoxicity. However,existing stilbene derivatives have not been available or practical aspharmaceutical agents for a number of reasons, including their lowsolubilities in water.

Recently, certain stilbene derivatives which inhibit tubulinpolymerization and having improved solubility in water have beeninvestigated, for instance, the phosphorylated pro-drugCombretastatin-A4, see U.S. Pat. No. 5,561,122. Other stilbenederivatives having improved solubility in water have been proposed asdrugs, such as those of the present Assignee as described for instancein Japanese Patent Kokai Publications JP-A-7-228558 and JP-A-8-301831.While such stilbene derivatives are promising, alone they may exertinsufficient anti-tumor activity. Thus, it is desired to enhance theantitumor efficacy and safety of compositions containing stilbenederivatives.

BRIEF SUMMARY OF THE INVENTION

It is an object of the present invention to develop a superior antitumoragent, specifically, to develop a pharmaceutical preparation whichimproves the efficacy of a stilbene derivative and, in particular, todevelop and provide an antitumor agent exhibiting superior safety andanti-tumor efficacy for reducing the severity, preventing, treating orcuring tumors and other neoplasms, particularly solid or highlymalignant tumors.

The present inventors have persistently and methodically conductedresearch with the above objectives in mind in order to overcome therecognized problems of the prior art compounds and therapies. Throughthese efforts, it has been discovered that a stilbene derivative whenadministered together with a platinum coordination compound, such asCisplatin, provide enhanced or synergistic anti-neoplastic or anti-tumoreffects and significantly enhance the tumor-inhibiting properties ofeither or both compounds.

Addition of a stilbene derivative to a therapeutic platinum coordinationcompound can provide superior anti-neoplastic or anti-tumor effectscompared to combinations of other chemotherapeutic drugs with a platinumcoordination compound, such as Cisplatin. Such stilbene-containingcompositions thus provide a chemotherapeutic drug higher in efficacyuseful to treat tumor or cancer patients. For instance, enhanced orsynergistic effects are achieved by employing a stilbene derivative incombination with platinum coordination compound, such as Cisplatin. Suchcompositions are used to more efficaciously and safely to treat or curetumors and solid cancers.

The present inventors have found that, by combining a stilbenederivative, most preferably one having in-vitro tubulin polymerizationinhibiting activity with a platinum coordination compound such asCisplatin, it is possible to completely cure tumors and preventdecreases in body weight. For instance, it is demonstrated that thepresent invention cures mice of malignant engrafted tumors. Thesestudies show that the inventive compositions are highly effectivechemotherapeutic agents against cancer and tumors, and thus may findsuccessful application for treating or curing various solid cancers ortumors, such as pulmonary or lung cancer.

Vincristine, Vindesine, and Vinblastine are examples of currently knowntherapeutic agents having tubulin polymerization inhibiting activities.These agents are used for treatment of solid cancers in multi-drug orpolypharmacy therapeutic approaches. They may be used in combinationwith drugs such as Cisplatin. However, complete curing of tumors in micehas not been observed after simultaneous administration of Vindesine andCisplatin. Further, the combination of Vindesine and Cisplatin is shownto result in significant decreases in body weight, and thus suchcombinations may be toxic and unsafe.

On the other hand, the inventive combination of Cisplatin and a stilbenederivative results in complete curing of tumors in mice and this effectis attributed to-the combined, enhanced or synergistic action of thecompositions of the present invention. Further, combinations of astilbene derivative and Cisplatin have improved safety as they do notcause significant decreases in weight. For instance, a compositioncomprising Cisplatin and the compound shown by the following structuralformula (3):

did not cause significant decreases in body weight compared to thecombination of Cisplatin with other anti-tumor agents, such asVindesine, see Table 3 below.

These results are compelling evidence of the highly efficaciousproperties of the inventive combination in treating tumors in animals,preferably mammals, and most preferably human cancer patients comparedto compositions containing Cisplatin and other (non-stilbene)chemotherapeutic drugs. Thus, the present invention provides an improvedanti-neoplastic or antitumor agent, especially for cancer chemotherapy,and provides a means for relieving patients from tumor burdens, as wellas safely reducing the side-effects of cancer therapy. That is, thepresent invention provides a high potency antitumor effect thusproviding a means for reducing the overall dosages of antitumor agentsadministered, and thus providing reductions in toxicity andside-effects.

Accordingly, the present invention using the combination of a stilbenederivative and a platinum coordination compound provides a novel andhighly efficacious compositions and methods for the treatment ofneoplastic diseases, especially tumors.

DETAILED DESCRIPTION OF THE INVENTION

As discussed above, the present invention is directed to an antitumoragent comprising al least one stilbene derivative and at least oneplatinum coordination compound. The stilbene derivative and platinumcoordination compound may be combined together in one composition, orseparately compounded and then administered together. For instance, theclaimed invention may comprise a kit comprising a stilbene derivativeand a platinum coordination compound as well as equipment and suppliesfor the administration of these compounds as well as instructions foruse.

The present invention also encompasses compositions comprising more thanone type of stilbene derivative or more than one type of platinumcoordination compound.

Combinations of stilbene derivatives and platinum coordination compoundswith other pharmacological agents, anti-tumor drugs or assistantanti-tumor agents are also encompassed by the invention.

The invention also encompasses methods of treatment involving thecoadministration or sequential administration of a stilbene derivativeand platinum coordination compound.

All sorts of neoplasms and tumors occurring in animals, especiallymammals, and most preferably humans, may be treated using thecompositions and methods of the present invention. Most preferably, theantitumor agent and methods of the present invention may be used forinhibiting the growth and proliferation of tumor cells in a human being.

There is no particular limitation to the form of administration of theantitumor agent. The platinum coordination compound can be routinelyadministered parenterally, and the stilbene derivative can also beadministered parenterally. However, the present invention alsoencompasses methods involving a combination of distinct methods orroutes of administration. For instance, the stilbene derivative can beadministered in a different form and route than the platinumcoordination compound, or according to a separate dosage schedule.

Preferably, the stilbene derivative used in the present invention hascis-stilbene as a fundamental skeleton and exhibits in vitro tubulinpolymerization inhibiting activity and/or an antitumor activity. Tumorcell proliferation inhibiting activity is the preferred type ofanti-tumor activity. Known compounds, as well as stilbene compoundswhich will be found in future, are included in the stilbene derivativesin the present invention provided that such newly found compounds areclassed as stilbene derivatives. The stilbene derivatives of the presentinvention also include bioprecursors or compounds which may be convertedin an animal body into a stilbene derivative. Any suitable orpharmaceutically allowable derivatives, such as salts, esters, solvates(salvation products) such as hydrates thereof, may be used as thestilbene derivatives in the present invention. Preferably, suchderivatives exhibit significant antitumor activity when used in vivo.The present compositions may also be used in vitro or in methods of exvivo treatment.

Among representative stilbene derivatives, having the cis-stilbene as afundamental skeleton, there are preferably compounds represented by thefollowing formulas (1) and (2):

and corresponding salts, hydrates and solvates (solvation products), andespecially pharmaceutically acceptable forms thereof.

In the above formulas, R¹, R² and R³ independently denote lower alkoxygroups, R⁴, R⁵ and R⁶ independently denote any substituent of a hydrogenatom, a halogen atom (fluorine, chlorine atoms, etc.), a nitro group, ahydroxyl group, a lower alkoxy group, a phosphoric acid ester (asubstituent formed on phosphoric acid esterification with a hydroxylgroup: —OPO₃H₂, hereinafter the same), a phosphoric acid amide (asubstituent formed on phosphoric acid amidation with an amino group:—NHPO₃H₂, hereinafter the same), an amino lower alkoxy group, a loweralkyl amino lower alkoxy group, a di-lower alkyl amino lower alkoxygroup, a mercapto group, a lower alkyl thio group, an amino group, alower alkyl amino group, a di-lower alkyl amino group, a lower alkylgroup, an amino lower alkyl group, a trifluoro methyl group, a loweralkanoyl group, a lower alkanoyl amino group and an amino acid acylaminogroup, X denotes a hydrogen atom or a nitrile group, and Het denotes aheterocyclic ring.

The number of carbon atoms in the above described lower alkyl group andthe lower alkoxy group is from 1 to 5 and the number of carbon atoms inthe lower alkanoyl group is from 2 to 6.

The amino acid acyl group in the amino acid acylamino group is an acylgroup derived from an amino acid. The amino acids may be α-amino acids,β-amino acids and γ-amino acids. Examples of preferred amino acidsinclude glycine, alanine, leucine, serine, lysine, glutamic acid,aspartic acid, threonine, valine, isoleucine, ornithine, glutamine,aspargine, tyrosine, phenylalanine, cysteine, methionine, arginine,β-alanine, tryptophan, proline, histadine, etc. The amino acids may beof the L-, D- or DL-form, preferably the L-form. In particular,threonine and serine are preferred in view of their pharmaceuticaleffects and safety.

The heterocyclic rings may be, for example, tetrazole ring, thiazolering and the like. If the heterocyclic ring is a thiazole ring, it mayhave a substituent of a lower alkyl group, an amino group, a mono-loweralkyl amino group, a di-lower alkyl amino group, a hydrazino group, ahalogen atom, such as fluorine and chlorine atom, and a lower alkoxygroup. The number of carbon atoms in the lower alkyl group and the loweralkoxy group is 1 to 5.

As described above, the stilbene derivative in the present invention isa compound having a cis-stilbene skeleton in its structure and whichexhibits a tubulin polymerization inhibiting activity and/or anantitumor activity. The stilbene derivative may be, for exampleCombretastatine-A4, or a tumor proliferation inhibitive stilbenederivative, disclosed in prior art publications, such as patentpublications, e.g. U.S. Pat. Nos. 4,996,237, 5,561,122 and 5,430,062,Japanese Patent Kokai Publications JP-A-7-228558, JP-A-8-301831 andJP-A-8-301831 and JP-A-10-81673, corresponding to Japanese patentApplication Serial No. 236603/1996 filed by the present Applicant onSep. 6, 1996. The prior art stilbene derivatives, described in thesepatent publications, can be used as the stilbene derivatives of thepresent invention, insofar as the prior art stilbene derivatives conformwith the above definition of the stilbene derivatives in the presentinvention. In addition, all the contents of the prior art patentpublications describing stilbene derivatives are incorporated herein byreference.

The above-mentioned stilbene derivatives may be manufactured by anyroutine technique including the methods disclosed in the above-mentionedpublications. It is noted that future stilbene derivatives may bemanufactured and used for the present invention in the same manner asdescribed above.

Among the stilbene derivatives of the present invention, there aresalts, esters, and other derivatives of stilbene, and derivatives whichmay be converted in vivo into stilbene derivatives, insofar as thestilbene derivatives manifest the above-mentioned objective activitiesin an animal body.

As the stilbene derivative used in the present invention, a compoundrepresented by the following formula (1) is preferred:

where R¹, R², R³ and R⁵ denote a methoxy group, R⁴ is an amino group oran amino acid acylamino group and R⁶ and X are hydrogen.

Particularly preferred among the compounds represented by the aboveformula (1), is a compound represented by the following formula (3),also referred to as Compound (3) below:

Compound (3) is(Z)-1-(3-amino-4-methoxyphenyl)-2-(3,4,5-trimethoxyphenyl)-ethene-L-serineamide, and is soluble in water. Compound (3) may be in the form of asalt, for example, hydrochloride, acetate, methanesulfonate and thelike.

The manufacture of Compound (3), which may be in the form of thepharmaceutically acceptable salts, hydrates and solvates, and themanufacture of oral and/or parenteral pharmaceutical compositionscontaining Compound (3), as well as inert pharmaceutically acceptablecarrier(s) and/or diluent(s), are extensively disclosed in JapanesePatent Kokai Publication JP-A-8-301831, which is specificallyincorporated by reference.

The platinum coordination compound used in the present invention is acompound which provides platinum, preferably in an ionic form, and alsoexhibits antitumor activity. More preferably, the platinum coordinationcompound exhibits tumor cell proliferation preventative (inhibiting)properties.

Specific examples of platinum coordination compounds, employed in thepresent invention, preferably include Cisplatin,cis-diamminediacoplatinum (II)-ion, chloro(diethylene-criamine)-platinum (II) chloride, dichloro(ethylenediamine)-platinum (II), diammine(1,1-cyclobutane dicarboxylate) platinum(II) (carboplatin), spiroplatin, iproplatin, diammine(2-ethylmalonate)-platinum (II), ethylene diammine malonate platinum(II), aqua (1,2-diaminodichlohexane)-sulfate platinum (II),(1,2-diaminocyclohexane) malonate platinum (II),(4-caloxyphthalate)-(1,2-diaminocyclohexane)-platinum (II),(1,2-diaminocyclohexane)-(isocitrate)-platinum (II),(1,2-diaminocyclohexane)-cis (pyruvate) platinum (II),(1,2-diaminocyclohexane)-oxalate platinum (II), Ormaplatin andtetraplatin.

Certain explanations will be hereinafter made as to a platinum complex,included in the platinum coordination compound according to the presentinvention, as a chemotherapeutic drug.

As the platinum coordination compounds used in the present invention,there are those compounds already known as the chemotherapeutic drugs,see Tamura, T. et al., Jpn J. Clin. Oncol. vol. 18 (1):27 (1988) andFukuda, M. et al., Cancer Chemother. Pharmacol., vol. 26: 393 (1990).

It is necessary to increase the efficacy of the tumor proliferationsuppressing activity of Cisplatin and other diamino-platinum complexesin order to reduce the tumor burden and help relieve the patient oftumor-associated pain.

Among the platinum coordination compounds used for the presentinvention, Cisplatin, Carboplatin and Nedaplatin are preferred due totheir curative effects. Although the compound “Cisplatin”, which meanscis-dichloro diamine platinum (II), may be manufactured by prior arttechniques, it is also available commercially. For example, Cisplatinmay be obtained, as a powder for constituting with water, an asepticphysiological saline water or other suitable excipient, under the tradename of Platinol (Registered Trademark) from Bristol Myers-Squibb Co. orunder the trade name of “randa Inj.” from NIPPON KAYAKU CO., LTD.

Other platinum coordination compounds used in the present invention arecommercially available or may be manufactured by known or routinely usedtechniques. Insofar as the platinum coordination compounds come withinthe definition therefor of the present invention, the platinumcoordination may be purchased or manufactured by manufacturing methodswhich will be developed in future.

When the antitumor agent of the present invention is to be used,stilbene derivatives in an amount sufficient to inhibit tumorproliferation may be combined with platinum coordination compounds, andadministered to a subject animal, preferably a mammal, and morepreferably a human being, in need of curing, alleviation or preventionof tumors, especially a human being suffering from tumor cellproliferation, to inhibit tumor cell proliferation.

As described above, the two types of such efficacious ingredients in thepresent invention may be combined in a pharmaceutical preparation withthe objective of obtaining an enhanced anti-tumor effect. For instance,a preferred embodiment of the present invention is to use compound (3)in an amount effective to inhibit tumor cell proliferation incombination with Cisplatin to inhibit tumor cell proliferation.

Moreover, a pharmaceutical preparation containing one of the twoefficacious ingredients is also encompassed by the present invention, ifsuch pharmaceutical preparation has the objective of being used incombination with the pharmaceutical preparation containing the otherefficacious ingredient in the present invention. This pharmaceuticalpreparation may contained, for example, as one component in atherapeutic kit.

The inhibition of tumor cell proliferation means inhibition ofproliferation of the tumor cells sensitive to therapy includingadministration of an effective amount of the stilbene derivatives, suchas compound (3), and the platinum coordination compounds, such asCisplatin, to, for example, a human being suffering from tumor cellproliferation. In an acceptable case, this administration suppressestumor cell proliferation or diminishes the measurable tumor size. In anoptimum case, the tumor undergoes complete regression.

As described above, there is no particular limitation to the method ofadministering the antitumor agent of the present invention to the humanbeing, such that it may be administered orally or parenterally, forinstance, intravenously, or by a subcutaneous or intramuscular route.For prompt efficacy, parenteral administration, such as by intravenousand subcutaneous administration, or by infusion, etc. is preferred.

In the method for administering the pharmaceutical preparation accordingto the present invention, the stilbene derivative may be administeredsimultaneously with the platinum coordination compound or the two may besequentially administered in an optional order. The practicallydesirable method and sequence for administration are varied depending onthe individual preparation of the stilbene derivative used, such as thecompound (3), individual preparation of the platinum coordinationcompound in use, such as Cisplatin, individual tumor cells being cured,and the individual subjects being treated. The optimum method andsequence for administration of the stilbene derivative and the platinumcoordination compound under preset given conditions may be suitablyselected by those skilled in the art with the aid of the routinetechnique and the information contained in the present specification.

Curative and tumor-inhibiting amounts of stilbene derivatives incombination with platinum coordination compounds may be calculated bythose with skill in the art based on the present disclosure. A curativeunit will inhibit proliferation of tumor cells sensitive to thesecompounds in the human being suffering from tumor cell proliferation.The practically desirable curative unit is varied depending on theindividual dosage forms of the stilbene derivative used, such ascompound (3), individual dosage forms of the platinum coordinationcompound, such as Cisplatin, individual tumor cells being cured and theindividual subjects being treated. The optimum curative units for presetgiven conditions may be suitably selected by those skilled in the artwith the aid of the curative test units and the information contained inthe present specification.

When administering the antitumor agent of the present invention, theadministration schedule for the platinum coordination compound ispreferably determined by setting approximately 1 to 500 mg/m² of thebody surface area per curative unit as a reference. When using Cisplatinand compound (3), these may be administered simultaneously, Cisplatinmay be administered either before or after the administration ofcompound (3); Alternatively, these compounds may be used in combination.The preferred amount of Cisplatin for administration is approximately 10to 100 mg/m² of the body surface area per day. It is preferablyadministered simultaneously along with curative units of Compound (3)continuously for one to five days at a time.

The platinum coordination compound in an infusion form is preferablyinfused once or twice a week. This weekly infusion is preferablyrepeated several times unless contraindicated by the appearance ofundesirable side effect actions such as nephrotoxicity andneurotoxicity. It is possible to use other conventionally usedtechniques simultaneously with the administration of the platinumcompound, such as Cisplatin or the other platinum coordination compound.

The antitumor agent of the present invention may comprise apharmaceutical preparation comprising at least the stilbene derivativeand the platinum coordination compound as described above, such that thetwo active ingredients may be contained as a mixture in thepharmaceutical preparation. However, the two active ingredients in thepresent invention may also be contained separately in distinctpharmaceutical preparations used in combination. It is noted that such apharmaceutical preparation containing other agents (third and fourthmedical ingredients and so on) such as other antitumor agents, maynaturally be encompassed by the present invention, insofar as theeffective ingredients used in the present invention are contained in thepharmaceutical preparation. Moreover, it is possible for carriers,diluents and other substances, pharmaceutically acceptable for any ofthe pharmaceutical preparations in the present invention (a solepharmaceutical preparation containing both ingredients in the presentinvention and separate pharmaceutical preparations separately eachcontaining one of the two ingredients for use in combination) to becontained in the antitumor agent of the present invention.

As the suitable pharmaceutically acceptable carriers and diluents, usedin the antitumor agent of the present invention, those carriers,diluents and excipients and the like known to those skilled in the artof preparation of pharmaceutical preparations, may be used asappropriate. Such carriers are disclosed in, for example, JapanesePatent Kokai Publication JP-A-8-301831 and the other aforementionedprior art publications.

The antitumor agent of the present invention may be suitably appliedparenterally, as discussed above. In this case, the antitumor agent isprepared into an intravenous infusion or injection, along withpharmaceutically acceptable carriers by various methods known to thoseskilled in the art. Preferably, the pharmaceutical agent is manufacturedby a routine technique e.g. in a unit dosage form and in the form of afreeze-dried mixture of two effective ingredients, which is re-preparedin water or another suitable liquid infusion prior to administration.

The ratio of the two ingredients for the pharmaceutical preparation forthe antitumor agent of the present invention may be varied in a widerange, depending on a number of factors, such as a desired amount ofadministration and on the pharmaceutically acceptable carrier in use.The ratios of the stilbene derivative and platinum coordination compoundof the present invention for use in a pharmaceutical preparation are forthe stilbene derivative approximately 0.01 to 1000 parts by weight,preferably approximately 0.1 to 100 parts by weight, to 1 part by weightof the platinum coordination compound. So, when the pharmaceuticalpreparation in the present invention containing two active ingredientsis to be administered to the patient, it is administered in an amountwhich will give the above-defined administration range.

If the pharmaceutical preparation is to be administered stepwise, theabove-defined administration range can be set as the average ratio forthe separate pharmaceutical preparations.

Preferably, 5 to 500 mg of the platinum coordination compound, morepreferably 10 to 50 mg as Cisplatin, 0.1 to 10,000 mg of the stilbenederivative and more preferably 1 to 1,000 mg as the compound (3) may becontained for each dosage of the pharmaceutical preparation according tothe present invention. It is desirable that mannitol and/or sodiumchloride be contained in routine amounts in the pharmaceuticalpreparation of Cisplatin. The physiological pharmaceutical value of thepharmaceutical composition used as an injection or infusion liquid issuitably adjusted by the content of a buffer well-known in the art.

Having generally described this invention, a further understanding canbe obtained by reference to certain specific examples which are providedherein for purposes of illustration only, and are not intended to belimiting unless otherwise specified.

Example 1 Antitumor Effect and Safety Tests

A pharmaceutical composition comprising the stilbene derivative offormula (3):

was prepared and used in the tests presented below:

Stilbene derivative of formula (3) as the hydrochloride 10 mgPhysiological saline solution in water: 10 ml

As Cisplatin, a pharmaceutical preparation marketed by NIPPON KAYAKUCO., LTD. under the trade name of “randa Inj.” (a preparation containing0.5 mg of Cisplatin in 1 ml of a solution) was used.

Methodology. 10 mg of a murine colonic tumor, colon 26, was inoculatedunder the skin of the back of CDF1 mice (day 0). After one week, thetumor was measured to calculate the volume of the tumor and the micewere classified into several groups (each group: n=5) and administrationof the stilbene derivative and Cisplatin was started. On days 7, 11 and15, the tumor-inoculated mice were bolusly injected under the skin oftheir backs and into their tail veins with 1 mg of the stilbenederivative of formula (3) in 1 ml of physiological saline solution and0.5 mg of Cisplatin (“randa Inj”) in a volume of 1 ml. Control andcomparative groups of mice were similarly injected with the other drugcombinations described in Table 1. The comparative drug Vindesine is anantitumor drug used in combination with Cisplatin by Garalla et al.VP-16 is also a chemotherapeutic drug which is often used clinically incombination with Cisplatin. The dosages of Cisplatin, VP-16 andVindesine represent the maximum dosage without death due to the toxicityin the administration schedule in the present example.

As indicated in Table 1 the term [−CDDP] means that Cisplatin was notadministered at all. The term [+CDDP] means that Cisplatin wasadministered in an amount of 5 mg/kg/day. The term [n.d.] means that notesting was performed.

Antitumor effects: Complete regression of the colonic tumor wasdetermined by palpation. Subjects in which the tumor has not beenascertained by palpation on the 60th day were deemed to have theirtumors cured completely. These results are shown in Table 1.

TABLE 1 Antitumor Activity Test (1): Number of instances Amount of ofcomplete regression Administration of tumors Sample mg/kg/day −CDDP+CDDP Control — 0/5 0/5 Compound (3) 5 n.d. 1/5 Compound (3) 10 0/5 4/5Vindesine 2 n.d. 0/5 VP-16 30 0/5 0/5

As clearly shown in the results of Table 1, the antitumor agent of thepresent invention, that is the combination of the stilbene derivativeand the platinum coordination compound, produced complete regression oftumors, as compared to a pharmaceutical preparation composed of any oneof two ingredients. This shows an enhanced or synergistic effect forthese two drugs when used together.

On the other hand, in the pharmaceutical preparation, used clinically incombination with Cisplatin, such as Vindesine or VP-16, the completetumor regression was not observed.

Safety. The rate of body weight change was used as a measure of safetyas significant loss of body weight is undesirable in cancer patients.Body weight change was calculated using the following equation:

Rate of body weight change (%)=[{(body weight−weight of the tumor) atthe 21^(st) day}−{(body weight−weight of the tumor) at the 7^(th)day}]/{body weight−weight of the tumor) at the 7^(th) day}×100.

Using the above equation, the rate of body weight change at the 21st daywas calculated. The results are shown in Table 2.

TABLE 2 Safety Test (1) Amount of Rate of Body Administration WeightChange Sample mg/kg/day −CDDP +CDDP Control — −12.0  −11.1 Compound (3)5 n.d. −4.2 Compound (3) 10 −7.7 −1.9 Vindesine 2 n.d. −23.2 VP-16 30−1.0 −9.9

Concerning the safety aspect, the high rates of body weight loss notedin the groups receiving the combination of Vindesine or VP-16 with theplatinum coordination compound, were not observed in test groupsreceiving the combination of the two ingredients of the presentinvention, as clearly shown in the results of Table 2.

Example 2 Antitumor Effect and Safety Tests (Preparation ofPharmaceutical Preparation)

Pharmaceutical preparations for infusion were prepared using stilbenederivatives (4) and (5), shown by the following chemical formulas:

The following compositions comprising these stilbene derivatives wereprepared:

Compound (4) (as the hydrochloride)   5 mg; Tween 80 0.5 ml; andPhysiological saline in water 9.5 ml. Compound (5) (as thehydrochloride)  10 mg; Tween 80 0.5 ml; and Physiological saline inwater 9.5 ml.

As in Example 1 as described above, the pharmaceutical preparation(“randa Inj.”) marketed by NIPPON KAYAKU CO., LTD. (containing 0.5 mg ofCisplatin in 1 ml solution) was used as the Cisplatin. Thepharmaceutical preparation (“paraplatin injection”) marketed by BristolMyers-Squibb Co. (containing 10 mg of Carboplatin in 1 ml) was used asthe Carboplatin.

Methodology: 10 mg of a colonic tumor of mouse, colon 26 was inoculatedunder the skin of the back of CDF1 mice (day 0). After one week, thetumor was measured to calculate the volume of the tumor and the micewere classified into several groups (each group: n=5) and administrationof the pharmaceutical preparation was started.

Stilbene derivatives of formulas (3), (4) and (5), and Cisplatin andCarboplatin were bolusly injected into the tail vein on the 7th, 11thand 15th days as described in Tables 3 through 6 below.

Antitumor effects: Complete regression of the colonic tumor wasdetermined by palpation. Subjects in which the tumor had not beenascertained by palpation on the 60th day were deemed to have theirtumors completely cured. These results are shown in Tables 3 and below.

The terms: [+CBDCA] and [+CDDP] indicate that 50 mg/kg/day ofcarboplatin or 5 mg/kg/day of cisplatin were administered, respectively.The terms: [−CBDCA] and [−CDDP] indicate that no carboplatin orcisplatin were administered. The dosages of cisplatin and carboplatindenote the maximum dosages without death due to toxicity in theadministration schedule in the present embodiment.

TABLE 3 Antitumor Activity Test (2) Number of instances Amount of ofcomplete regression Administration of tumors Sample mg/kg/day −CBDCA+CBDCA Control — 0/6 0/6 Compound (3) 20 0/6 1/6

TABLE 4 Antitumor Activity Test (3) Number of instances Amount of ofcomplete regression Administration of tumors Sample mg/kg/day −CDDP+CDDP Control — 0/6 0/6 Compound (4)  5 0/6 4/6 Compound (5) 20 0/6 6/6

As clearly shown in Tables 3 and 4, the antitumor agent of the presentinvention comprising a stilbene derivative exhibits superior antitumoractivity in combination with a platinum coordination compound.

Safety: The rate of body weight change was used as a measure of safety,as significant loss of body weight is undesirable in cancer patients.Body weight change on the 21^(st) day was calculated using the equationdescribed in Example 1. These results are shown in tables 5 and 6.

TABLE 5 Safety Tests (2) Amount of Rate of Body Administration WeightChange Sample mg/kg/day −CBDCA +CBDCA Control — −22.1 −9.6 Compound (3)20 −7.4 1.8

TABLE 6 Safety Test (3) Amount of Rate of Body Administration WeightChange Sample mg/kg/day −CDDP +CDDP Control — −22.1 −11.4 Compound (4) 5 −7.0 −13.9 Compound (5) 20 −2.1 −0.3

Moreover, concerning the safety aspect, as clearly shown in the resultsof Tables 5 and 6, outstanding improvement may be noticed by employingthe two ingredients of the present invention.

Finally, it may be seen from the results of Tables 1 to 6 that, with thecombination of the stilbene derivative and the platinum coordinationcompound according to the present invention, the efficacy as theantitumor agent can be improved synergistically beyond the expectationby those skilled in the art, such that a practically highly usefulantitumor effect can be achieved.

EFFECTS OF THE INVENTION

The anti-tumor agent according to the present invention, comprising astilbene derivative and a platinum coordination compound, such asCisplatin, in combination or as a mixture, can be used as an anti-tumoragent, especially as a chemotherapeutic drug or agent for cancertreatment. The inventive combination of a stilbene derivative with aplatinum coordination compound is highly effective for treatment,suppression, prevention or cure of tumors, especially solid cancers ortumors, such as solid carcinoma. Some of these anti-tumor effects may beattributed to a synergistic effect derived from the combination of thestilbene derivative and platinum coordination compound.

Modifications and Other Embodiments

Various modifications and variations of the described anti-tumor agents,compositions and methods as well as the concept of the invention will beapparent to those skilled in the art without departing from the scopeand spirit of the invention. Although the invention has been describedin connection with specific preferred embodiments, it should beunderstood that the invention as claimed is not intended to be limitedto such specific embodiments. Various modifications of the describedmodes for carrying out the invention which are obvious to those skilledin the medical, biological, chemical or pharmacological arts or relatedfields are intended to be within the scope of the following claims.

INCORPORATION BY REFERENCE

Each document, patent application or patent publication cited by orreferred to in this disclosure is incorporated by reference in itsentirety. Any patent document to which this application claims priorityis also incorporated by reference in its entirety. Specifically,priority documents JP 10/108,708, filed Apr. 3, 1998 and JP 10/229,843,filed Aug. 14, 1998 are hereby incorporated by reference.

1. A composition, comprising synergistic effective amounts of: (a) astilbene compound of formula (1), or a pharmaceutically acceptable saltthereof, a compound of formula (2), or a pharmaceutically acceptablesalt thereof:

wherein R¹, R² and R³ are each independently a lower alkoxy group; R⁴,R⁵ and R⁶ are each independently selected from the group consisting of:hydrogen, a halogen, a nitro group, a hydroxyl group, a lower alkoxygroup, a phosphoric acid ester, a phosphoric acid amide, an amino loweralkoxy group, a lower alkyl amino lower alkoxy group, a di-lower alkylamino lower alkoxy group, a mercapto group, a lower alkyl thio group, anamino group, a lower alkyl amino group, a di-lower alkyl amino group, alower alkyl group, an amino lower alkyl group, a trifluoromethyl group,a lower alkanoyl group, a lower alkanoyl amino group, and an amino acidacylamino group; X is a hydrogen atom or a nitrile group; and Het is aheterocyclic ring; and (b) a platinum coordination compound selectedfrom the group consisting of cis-diaminedichloroplatinum (II),cis-diamminediaquoplatinum (II)-ion, chloro(diethylenetriamine)-platinum (II) chloride, dichloro(ethylenediamine)platinum (II), cis-diammine(1,1-cyclobutanedicarboxylato) platinum (II),Spiroplatin, Iproplatin, diammine (2-ethylmalonato)-platinum (II),ethylenediamminemalonatoplatinum (II),aqua(1,2-diaminocyclohexane)sulfatoplatinum (II),(1,2-diaminocyclohexane) malonatoplatinum (II),(4-caroxyphthalato)(1,2-diaminocyclohexane)-platinum (II),(1,2-diaminocyclohexane)(isocitrato)platinum (II),(1,2-diaminocyclohexane)cis (pyruvato)platinum (II),(1,2-diaminocyclohexane)oxalateplatinum (II), Ormaplatin, Tetraplatin,and Nedaplatin; wherein: a combination of cis-diaminedichloroplatinum(II) and a stilbene compound represented by formula (1) wherein R¹, R²,R³ and R⁵ are a lower alkoxy group, R⁴ or R⁶ is a hydroxyl group or ahydrogen such that when R⁴ is a hydroxyl group R⁶ is a hydrogen and whenR⁴ is a hydrogen R⁶ is a hydroxyl group and X is a hydrogen is excluded;and a combination of cis-diaminedichloroplatinum (II) and a stilbenecompound represented by formula (3):

is excluded.
 2. The composition of claim 1, wherein said stilbenecompound is represented by formula (1):

or a salt thereof; wherein R¹, R², R³ and R⁵ are each a methoxy group;R⁴ is an amino group or an amino acid acylamino group; and R⁶ and X arehydrogen.
 3. The composition of claim 1, wherein said stilbene compoundis:

or a pharmaceutically acceptable salt thereof.
 4. The composition ofclaim 1, wherein said stilbene compound is:

or a pharmaceutically acceptable salt thereof.
 5. The composition ofclaim 1, wherein the stilbene compound is

or a pharmaceutically acceptable salt thereof and said platinumcoordination compound is diammine(1,1-cyclobutanedicarboxylato)platinum(II).
 6. The composition of claim 1, wherein the stilbene compound is

or a pharmaceutically acceptable salt thereof and said platinumcoordination compound is diammine(1,1-cyclobutanedicarboxylato)platinum(II).
 7. The composition of claim 1, wherein the stilbene compound is

or a pharmaceutically acceptable salt thereof and said platinumcoordination compound is (1,2-diaminocyclohexane)oxalateplatinum (II).8. The composition of claim 1, wherein the stilbene compound is

or a pharmaceutically acceptable salt thereof and said platinumcoordination compound is (1,2-diaminocyclohexane)oxalateplatinum (II).9. The composition of claim 1, wherein the stilbene compound is

or a pharmaceutically acceptable salt thereof and said platinumcoordination compound is (1,2-diaminocyclohexane)oxalateplatinum (II).10. The composition of claim 1, wherein Het is a tetrazole ring or athiazole ring, said thiazole ring optionally substituted with a loweralkyl group, an amino group, a mono-lower alkyl amino group, a di-loweralkyl amino group, a hydrazino group, a halogen atom, or a lower alkoxygroup.
 11. The composition of claim 1, wherein said (a) stilbenecompound is present in a ratio of from 0.01 to 1,000 parts by weight per1 part by weight of said (b) platinum coordination compound.
 12. Thecomposition of claim 1, wherein said (a) stilbene compound is present ina ratio of from 0.1 to 100 parts by weight per 1 part by weight of said(b) platinum coordination compound.
 13. An anti-tumor agent, comprisingthe composition according to claim
 12. 14. A method for treating aneoplasm, cancer, a malignant tumor or a solid tumor in a patient,comprising administering an effective amount of the anti-tumor agentaccording to claim 13 to a patient in need thereof.
 15. An anti-tumoragent, comprising the composition according to claim
 1. 16. A method fortreating a neoplasm, cancer, a malignant tumor or a solid tumor in apatient, comprising administering an effective amount of the anti-tumoragent according to claim 15 to a patient in need thereof.